Related compounds[CautionAvoid contact with o-tolidine when performing this test, and conduct the test in a well-ventilated hood.]

Detection reagent
Dissolve 200 mg of o-tolidine in 2.0 mL of glacial acetic acid with the aid of a hot water bath. Dilute with water to 100.0 mL, and filter. Mix one volume of this solution with an equal volume of potassium iodide solution (0.83 in 100).

Procedure
Apply separately 10 µL of the Test preparation, Identification preparation, Standard preparation A, and Standard preparation B to a suitable thin-layer chromatographic plate (see Chromatography 621, coated with a 0.25-mm layer of chromatographic silica gel mixture. Place the plate in a chromatographic chamber, and develop the chromatograms in a solvent system consisting of a mixture of butyl alcohol, glacial acetic acid, and water (4:1:1) until the solvent front has moved about three-fourths of the length of the plate. Remove the plate from the developing chamber, and dry under a current of warm air. Transfer the dry plate to another chromatographic chamber containing a beaker with 1 g of potassium permanganate. Add 10 mL of dilute hydrochloric acid (4 in 10) to the beaker. Cover the chamber, and allow the chamber to become saturated with chlorine gas. Expose the plate to the chlorine gas for 8 minutes. Remove the plate and expose to the air for 2 minutes, then spray with freshly prepared Detection reagent: the intensity of the secondary spot from the Test preparation, corresponding in RF value to the principal spot from the Identification preparation, is not greater than that of the principal spot from the Identification preparation (1.0%), and no other secondary spot from the Test preparation is greater in intensity than the principal spot from Standard preparation A (0.5%). The sum of all secondary spots from the Test preparation is not more than 2.0%.